血管外膜成纤维细胞表达基质金属蛋白酶和组织基质金属蛋白酶抑止物的研究

Vascular adventitial fibroblasts express matrix metalloproteinases and tissue inhibitors of matrix metalloproteinases

目的血管外膜成纤维细胞能表达分泌细胞外基质如胶原蛋白从而参与血管重塑过程。本研究观察血管紧张素2诱导体外培养的血管外膜成纤维细胞过程中基质金属蛋白酶MMP和组织基质金属蛋白酶抑止物TIMP的表达,以揭示血管外膜成纤维细胞表达分泌胶原蛋白的分子机制。方法使用贴壁法体外培养大鼠胸主动脉外膜成纤维细胞;用实时RT-PCR技术检测血管紧张素2诱导后2h、6h、12h、24h的MMP2、MMP9、TIMP1、TIMP2、TIMP3mRNA在血管外膜成纤维细胞的表达。使用免疫印迹法检测血管紧张素2诱导后MMP2和TIMP1蛋白在血管外膜成纤维细胞的表达。结果 MMP2、TIMP1、TIMP2、TIMP3 mRNA在血管外膜成纤维细胞表达,而MMP9 mRNA在血管外膜成纤维细胞不表达。MMP2 mRNA在血管紧张素2诱导后表达下调,诱导24h下调约50%。TIMP1mRNA在血管紧张素2诱导后表达上调,诱导12h上调约60%。TIMP2和TIMP3的mRNA表达在血管紧张素2诱导后没有明显变化。MMP2蛋白在血管紧张素2诱导后表达下调,诱导24h下调约50%。TIMP1的蛋白表达在血管紧张素2诱导后表达上调,诱导12h上调约50%。结论血管紧张素2可以调节血管外膜成纤维细胞MMP2和TIMP1的表达,MMP2和TIMP1表达变化可能参与血管外膜成纤维细胞表达分泌胶原。

Objective Adventitial fibroblasts differentiation to myofibroblasts plays an important role in vascular remodeling. Myofibroblasts proliferate, migrate and secrete extracellular matrix such as collagen. But the molecular events underling the collagen expression from adventitial fibroblasts remain unclear. The aim of this study was to investigate the expression of matrix metalloproteinases and tissue inhibitor of matrix metalloproteinases in cultured adventitial fibroblasts response to angiotensinⅡ(AngⅡ) stimulation. Methods Vascular adventitial fibroblasts were isolated and cultured from thoracic aorta of Sprague-Dawley rat. MMP2, MMP9, TIMP1, TIMP2 and TIMP3 mRNA expression were assayed by Real-time RT-PCR. MMP2 and TIMP1 protein expression were assayed by Western blot. Results MMP2, TIMP1, TIMP2 and TIMP3 but not MMP9 mRNA were expressed in cultured AF. AngⅡ time-dependently down-regulated MMP2 mRNA expression by 50% and up-regulated TIMP1 mRNA expression by 60%, respectively. TIMP2 and TIMP3 mRNA expression did not change upon AngⅡ stimulation. Consistently, AngⅡ downregulated MMP2 protein expression by 50% and up-regulated TIMP1 protein expression by 50%. Conclusion AngiotensinⅡ could regulate MMP2 and TIMP1 expression in cultured adventitial fibroblasts, which may contribute to collagen expression in adventitial fibroblasts.